title: Enhancing (L)-isoleucine production by thrABC overexpression combined with alaT deletion in Corynebacterium glutamicum.
abstract: L-isoleucine is synthesized from 2-ketobutyrate and pyruvate in Corynebacterium glutamicum, and the supplies of these two precursors are important for L-isoleucine synthesis. C. glutamicum YILWΔalaT with alaT gene deletion (encoding alanine aminotransferase, a principal enzyme for L-alanine synthesis) was constructed to increase intracellular pyruvate availability, and the thrABC genes from Escherichia coli (encoding bifunctional aspartate kinase I-homoserine dehydrogenase I, homoserine kinase, and threonine synthetase) were overexpressed in C. glutamicum YILW and YILWΔalaT to increase the supply of intracellular 2-ketobutyrate. In the fed-batch fermentation, YILWpXMJ19thrABC, YILWΔalaT, and YILWΔalaTpXMJ19thrABC exhibited 5.3, 17.6, and 8.4 % higher L-isoleucine production than the original strain, respectively. Both YILWpXMJ19thrABC and YILWΔalaT excreted lower concentrations of L-lysine, L-alanine, and L-valine. YILWΔalaTpXMJ19thrABC exhibited a cumulative reduction of these by-products excretion, which indicated that thrABC overexpression combined with alaT deletion resulted in the metabolic flux redistribution from 2-ketobutyrate and pyruvate to L-isoleucine synthesis, and decreased the fluxes to by-products synthesis accordingly.
authors.affiliation: National and Local United Engineering Lab of Metabolic Control Fermentation Technology, Tianjin University of Science and Technology, Tianjin, 300457, China.
authors.forename: Jing
authors.initials: J
authors.lastname: Wang
authors.forename: Bing
authors.initials: B
authors.lastname: Wen
authors.forename: Jian
authors.initials: J
authors.lastname: Wang
authors.forename: Qingyang
authors.initials: Q
authors.lastname: Xu
authors.forename: Chenglin
authors.initials: C
authors.lastname: Zhang
authors.forename: Ning
authors.initials: N
authors.lastname: Chen
authors.forename: Xixian
authors.initials: X
authors.lastname: Xie
mesh_terms: D000410:Alanine Transaminase / Q000172:deficiency* / Q000235:genetics*; D002087:Butyrates / Q000378:metabolism; D048230:Corynebacterium glutamicum / Q000166:cytology / Q000201:enzymology / Q000235:genetics* / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000235:genetics*; D005285:Fermentation; D017353:Gene Deletion; D015870:Gene Expression; D005818:Genetic Engineering / Q000379:methods*; D042541:Intracellular Space / Q000378:metabolism; D007532:Isoleucine / Q000096:biosynthesis*; D019289:Pyruvic Acid / Q000378:metabolism
chemical_list: D002087:Butyrates; D029968:Escherichia coli Proteins; D007532:Isoleucine; C005087:alpha-ketobutyric acid; D019289:Pyruvic Acid; D000410:Alanine Transaminase